酶放大镧系元素发光时间分辨荧光分析中 关键试剂的研究 (pdf)

    【摘要】 目的 研制成镧系元素发光时间分辨荧光分析中的关键试剂。 方法 用过碘酸钠法研制成辣根过氧化物酶标记链霉亲和素，用戊二醛二步法研制成碱性磷酸酶标记链霉亲和素和用化学合成法研制成5-氟水杨酸磷酸酯等关键试剂。结果 HRP标记SA总蛋白回收率为29.8%，纯度为97%。ALP标记SA的总蛋白回收率为56%，纯度为98%，分子量为159kD，其中ALP分子量为94kD，生物活性良好。5-氟水杨酸磷酸酯，测得熔点为157℃～159℃、纯度为99.43%～100%、红外吸收光谱、紫外吸收光谱及核磁共振谱等特性鉴定与文献记载一致。结论 研制成的各种关键试剂质量良好，已成功地应用于科研中。

    【关键词】 酶放大镧系元素发光；辣根过氧化物酶标记链霉亲和素；碱性磷酸酶标记链霉亲和素；5-氟水杨酸磷酸酯

    The studies of the key reagents for enzyme-amplified lanthanide luminescence time-resolved fluorescence assay

    SONG Na-ling，ZHAO Qi-ren，LI Mei-jia，et al. The Key Laboratory of Molecular Nuclear Medicine, Institute of Radiation Medicine, Chinese Academy of Medical Sciences and Peking Union Medical College, Tianjin 300192,China

    【Abstract】 Objective The key reagents were studied for enzyme-amplified lanthanide luminescence (EALL) time-resolved fluorescence assay (TRFA). Methods Horseradish peroxidase-labeled streptavidin (HRP-SA) was developed by sodium periodate method, alkaline phosphatase-labeled streptavidin (ALP-SA) was developed by two steps method of glutaraldehyde and 5-fluorosalicyl phosphate(5-FSAP) was synthesized chemically. Results The recovery rate of total protein of HRP-SA was 29.8%, purity was 97%. The recovery rate of total protein of ALP-SA was 56%, purity was 98%, the molecular weight was 94kD, biological activity was good. Determined 5-FASP melting point was 157℃～159℃, purity was 99.43%～100%, the characters of infrared absorption spectrum, ultra-violet absorption spectrum and nuclear magnetic resonance spectrum accorded with literature recordation. Conclusion The developed key reagents had good quality and were used successfully in EALL FRFA. ......